NHGRI/DIR Flow Cytometry Core

National Human Genome Research Institute

Investigators

Linked publications, trials & patents

Abstract

The NHGRI Flow Cytometry Core currently has three flow cytometers that are primarily used for sorting cells: two Becton Dickinson FACSArias, and one BD FACSVantage, SE Diva sorter. Each FACSAria is configured with three lasers and can measure up to nine fluorescent parameters as well as physical parameters (size and granularity). The Aria can sort 20-30,000 cells/second with high purity and recovery. Cells can be sorted into 4 populations simultaneously. Among others, over the past year, these sorting capabilities have been used for isolation tissue cell populations for animal transplantation experiments; isolation of blood cell sub-populations for analysis of functional cellular properties and gene transcription profiles, high throughput screening of enhancer regions. The FACSArias typically have a two to three week sign up window and are currently used at maximum capacity. A Fluidics Upgrade is being purchased for one of the FACSArias. Such upgrade that BD offers is a new fluidics system that is designed to achieve easier aseptic sort setup. There was high interest for this upgrade among the users of GMBB and GDRB that contributed FY08 funds for its purchase.[unreadable] The FACSVantage SE Diva sorter is equipped with three lasers and is used for applications different from the FACSAria. The FACSVantage can sort up to 4-6,000 cells per second and can sort four ways. This sorter is currently used for applications that require low pressure or UV laser excitation. These applications include the functional isolation of murine stem cells and dissection of zebra fish mesoderm and neurectoderm precursors cells. BD has informed us that they will no longer offer service contracts on the FACSVantage in 2010. [unreadable] For applications not involving cell sorting, but direct flow cytometry analysis of a variety of cell types, the Flow Core offers two BD FACSCaliburs, and one BD LSRII. The FACSCaliburs can measure up to four fluorescent parameters, and are routinely used for data acquisition and pre-sort analysis. Typical uses include analysis of GFP expression and cell cycle, as well as mutagenesis screening. The use of the FACSCaliburs has decreased as more investigators move to the LSRII for their analysis. We anticipate in the near future that we will need to replace one of the FACSCaliburs whose technology is becoming obsolete with either a second LSRII or a FACSCanto. Both of the latter instruments are digital, multi-color analyzers. [unreadable] The LSRII uses digital electronics and Diva software similar to the Aria. The LSRII is used to perform 9-color analysis. This allows investigators to characterize cells in more detail before sorting on the Aria. The LSRII use is currently at maximum capacity. The LSRII is routinely used during regular Core hours and after hours. Several investigators have begun using the High Throughput Sample (HTS) device, which has given them the ability to analyze many samples in 96 well plate format without the need of sitting at the instrument during the analysis procedure. [unreadable] The Flow Core also maintains a Miltenyi Auto MACS that is used for magnetic cell separations. The AutoMACS is often used as a pre-enrichment step prior to sorting on a flow cytometer. [unreadable] Over the past year, the services and capabilities of the NHGRI Flow Core have been taken advantage of by over 60 trainees from 5 Branches/14 Sections in the Institute (Table 1).[unreadable] For FY08, the Flow Core received a budget of $244,611. Of that total amount over $146,000 was allotted for service contracts on the instruments. This budget did not include any extra money for the acquisition of new equipment. No budget surplus is anticipated at the end of FY08[unreadable] During the past year, the Flow Core continued to maintain CLIA accreditation in support of immunophenotype and protein expression studies used in NHGRI clinical research protocols.[unreadable] [unreadable] 2008 NHGRI Flow Cytometry Core Use from September 2007 - June 2008[unreadable] [unreadable] Number of Branches 5[unreadable] Number of investigators 60[unreadable] Recorded Visits 1330[unreadable] Sorts 342